Fifteen years follow up with the St. Jude Medical Biocor porcine bioprosthesis.

BACKGROUND AND AIM OF STUDY: The study aim was to review the 15-year results of aortic (AVR) and mitral (MVR) valve replacement with the St. Jude Medical Biocor porcine prosthesis, in order to investigate long-term survival and valve-related complications. METHODS: Between January 1983 and January 1998, a total of 1,187 patients underwent either AVR (n = 1,029; mean age 69 years) or MVR (n = 158; mean age 63 years). Follow up (99.7% complete) was monitored in 1998, and all data were analyzed with regard to actuarial valve failure rates. Long-term echocardiographic data were obtained. RESULTS: Cumulative follow up time was 5,049 patient-years (pt-yr) for AVR patients, and 845 pt-yr for MVR patients. Actuarial survival rate at 15 years was 41 +/- 3%, and freedom from valve-related death was 94 +/- 1% for the AVR group; corresponding values for the MVR group were 25 +/- 11% and 84 +/- 6%. The occurrence of structural valve deterioration (SVD) varied with age; older patients were less affected. Freedom from SVD was 76 +/- 7% and 92 +/- 4% for AVR and MVR patients, respectively. Thromboembolism (TE) occurred mainly among the oldest patients, and was most prevalent among those with MVR. Actuarial freedom from TE was 82 +/- 5% after AVR and 75 +/- 7% after MVR. Prosthetic valve endocarditis (PVE) was rare, but caused the only reoperative mortality. Freedom from PVE was 95 +/- 2% after AVR and 93 +/- 3% after MVR. CONCLUSION: Although the optimal valve substitute remains to be found, this long-term study of a third-generation bioprosthesis showed a low incidence of valve-related complications, especially of valve deterioration. This type of bioprosthesis appears to be more durable than valves of previous generations.

Corticotropin-releasing factor--immunolabeled fibers in brain regions with localized kainate neurotoxicity.

Rats treated with the neuroepileptic drug, kainic acid, exhibit a specific regional pattern of neurodegeneration 24 h following onset of acute limbic status epilepticus. At 24 h post-seizure, the areas undergoing neurodegeneration also exhibit substantial amounts of the neuropeptide corticotropin-releasing factor (CRF) which is not present under normal conditions. In experimental brains, CRF is localized immunocytochemically to cells and densely labeled fibers in areas with neurodegeneration. Networks of CRF fibers closely surround moribund neurons staining intensely for acid fuchsin. Acid fuchsin, an acidophilic dye, is used routinely as a marker for irreversible neuronal injury, and acid fuchsin-positive neurons are identified in specific areas affected by kainic neurotoxicity. Evidence exists in the literature that CRF functions in brain as a excitatory neurotransmitter/neuromodulator. Under certain pathological conditions (i.e., seizures, brain trauma, ischemia), it has been postulated that CRF could act as an neurotoxic agent. This study provides anatomical evidence that CRF may function following seizures as an neurotoxin because of the close proximity of CRF-labeled fibers to degenerating neurons.

Design of an adenosine phosphorylase by active-site modification of murine purine nucleoside phosphorylase. Enzyme kinetics and molecular dynamics simulation of Asn-243 and Lys-244 substitutions of purine nucleoside phosphorylase.

Our objective was to alter the substrate specificity of purine nucleoside phosphorylase such that it would catalyse the phosphorolysis of 6-aminopurine nucleosides. We modified both Asn-243 and Lys-244 in order to promote the acceptance of the C6-amino group of adenosine. The Asn-243-Asp substitution resulted in an 8-fold increase in K(m) for inosine from 58 to 484 microM and a 1000-fold decrease in k(cat)/K(m). The Asn-243-Asp construct catalysed the phosphorolysis of adenosine with a K(m) of 45 microM and a k(cat)/K(m) 8-fold that with inosine. The Lys-244-Gln construct showed only marginal reduction in k(cat)/K(m), 83% of wild type, but had no activity with adenosine. The Asn-243-Asp;Lys-244-Gln construct had a 14-fold increase in K(m) with inosine and 7-fold decrease in k(cat)/K(m) as compared to wild type. This double substitution catalysed the phosphorolysis of adenosine with a K(m) of 42 microM and a k(cat)/K(m) twice that of the single Asn-243-Asp substitution. Molecular dynamics simulation of the engineered proteins with adenine as substrate revealed favourable hydrogen bond distances between N7 of the purine ring and the Asp-243 carboxylate at 2.93 and 2.88 A, for Asn-243-Asp and the Asn-243-Asp;Lys-244-Gln constructs respectively. Simulation also supported a favourable hydrogen bond distance between the purine C6-amino group and Asp-243 at 2.83 and 2.88 A for each construct respectively. The Asn-243-Thr substitution did not yield activity with adenosine and simulation gave unfavourable hydrogen bond distances between Thr-243 and both the C6-amino group and N7 of the purine ring. The substitutions were not in the region of phosphate binding and the apparent S(0.5) for phosphate with wild type and the Asn-243-Asp enzymes were 1.35+/-0.01 and 1.84+/-0.06 mM, respectively. Both proteins exhibited positive co-operativity with phosphate giving Hill coefficients of 7.9 and 3.8 respectively.

Reproducible fentanyl doses delivered intermittently at different time intervals from an electrotransport system.

The electrotransport transdermal fentanyl system (ET [fentanyl]), uses a small electrical current to enhance delivery of fentanyl to systemic circulation. Intermittent doses can be administered by periodic application of the current. The purpose of this study was to compare the effects of the frequency of intermittent drug delivery by ET (fentanyl) and compare the drug delivery to systemic circulation by ET (fentanyl) with intravenous administration. The topical safety was also determined for the ET (fentanyl) system. Nine adult male volunteers completed this three-treatment, randomized, 24-h, crossover study. ET (fentanyl) treatments with 200 microA direct current applied for 30 min at frequent (hourly) or infrequent (4-hourly) intervals over a 24-h period were compared. Also, the drug delivery to systemic circulation from ET (fentanyl) was compared with intravenous fentanyl 75 microg infused over 30 min every 4 h over a 24-hour period. The mean serum fentanyl concentration achieved with the hourly ET (fentanyl) regimen was higher than that for the 4-hourly ET (fentanyl) regimen as expected from the higher frequency of drug doses. The amount of fentanyl delivered estimated per dose from the ET (fentanyl) system using the iv fentanyl treatment as the reference was similar for the two ET regimens throughout the dosing period. This indicates consistent drug delivery regardless of the frequency of ET dosing. The majority of subjects reported either no, or barely perceptible, erythema 24 h after removal of the system.

Increased corticotropin-releasing factor immunoreactivity in select brain sites following kainate elicited seizures.

The literature has focused on the localization, regulation and function of corticotropin-releasing factor (CRF) expressing neurons localized in the paraventricular nucleus (PVN) of hypothalamus. However, less information is available on the expression, regulation, and function of CRF at extrahypothalamic sites. The current study examined the induction of CRF in extrahypothalamic brain sites following generalized clonic seizures induced by kainic acid. At 24 h post seizure onset, there was a marked increase of CRF immunolabeled perikarya in select brain areas, which contained little, if any, CRF in control brains. This CRF-like labeling was observed in olfactory structures such as the main olfactory bulb (internal granular layer), anterior olfactory nucleus, and deep layers of piriform cortex. Other sites of increased CRF-like immunoreactivity included the tenia tecta, inner layers of cingulate cortex, lateral septum, dorsal endopiriform nucleus, fundus striatum, and nucleus of the lateral olfactory tract. Additionally, CRF-like labeling was atypically increased in the amygdala (lateral and basolateral amygdaloid nuclei) and hippocampal formation (pyramidal cells of regions CA1/CA3 and polymorph cells within the dentate hilus). An association between the increased CRF immunoreactivity and neuropathological processes, characteristic of this seizure model, is hypothesized and discussed.

Effects of generalized convulsive seizures on corticotropin-releasing factor neuronal systems.

Most stressors generate a set of endocrine and neural adaptations that form a stress response. The corticotropin-releasing factor neurons of the paraventricular nucleus of hypothalamus integrate endocrine and neural inputs, and cause a cascade of events with resultant increased levels of pituitary adrenocorticotropic hormone and adrenal hormones. Although activation of the hypothalamic-pituitary-adrenal axis is associated with a large variety of stressors, the effects of seizures on hypothalamic corticotropin-releasing factor neurons are essentially unknown. The goal of the present study was to elucidate the effects of generalized convulsive seizures on distinct and separate corticotropin-releasing factor cell populations in brain. Seizure-activated neurons were identified immunocytochemically through their expression of the Fos protein. Seizures were induced by intraperitoneal injection of kainic acid. In the paraventricular nucleus, the vast majority of corticotropin-releasing factor-like parvocellular neurons also expressed Fos-like protein following seizure elicitation. This response was specific to corticotropin-releasing factor neurons of the paraventricular nucleus, as corticotropin-releasing factor neurons in central nucleus of the amygdala or bed nucleus of the stria terminalis did not simultaneously localize Fos following seizures.

Thrombophilia and lipid profile in post-menopausal women using a new transdermal oestradiol patch.

OBJECTIVE: To assess the changes, over a 6-month period, in serum lipoproteins, apoproteins and coagulation factors, induced in post-menopausal women treated by a new transdermal oestradiol patch. METHODS: Fifty-three hysterectomised, healthy, post-menopausal women were treated by a new transdermal patch designed to deliver 50 micrograms of 17 beta oestradiol per day (Gynaderm, Shire Developments). One patch was applied twice weekly. RESULTS: Forty-two patients completed the study. There was no significant change in the level of total cholesterol, triglycerides, HDL or LDL. There was a significant rise in the level of ApoAI after 3 months on therapy but this was not sustained after 6 months; there was also a significant drop in the level of ApoAII after 6 months on treatment. The changes in ApoB and Lp(a) were not statistically significant. There was a significant drop in the level of antithrombin III and of protein S, and a significant rise in factor VII. The drop in the level of fibrinogen and of protein C were not statistically significant. CONCLUSION: The transdermal route of oestradiol administration causes minimal changes in lipoprotein metabolism. The statistically significant changes in the thrombophilia profile parallel those observed with oral HRT, but, similarly, may not reflect clinical significance. The potential of transdermal oestrogens as cardioprotective agents is yet to be determined.

Corticotropin-releasing factor induction in rat piriform cortex following kainate-elicited seizures.

There is a substantial increase in number of cells labeled for corticotropin-releasing factor-like immunoreactivity in specific extrahypothalamic brain regions, particularly the piriform cortex, in rats allowed to survive 24 h following generalized clonic seizures. Seizures were elicited by kainic acid. Vehicle treated control animals had only a few cells labeled for corticotropin-releasing factor-like immunoreactivity at these brain sites. These areas of corticotropin-releasing factor-like induction appear to be localized to brain regions known to be vulnerable to kainate neurotoxicity and cell destruction.

A breast feeding education and promotion program: effects on knowledge, attitudes, and support for breast feeding.

This study was undertaken to determine the effects of a partner-support, incentive-based educational program on breast feeding knowledge, attitudes and support and to examine the relationship between feeding intentions and feeding behavior among low-income women. Women who expressed a willingness to participate in the intervention were randomly assigned to "intervention" and "usual breast feeding" (control) groups. Sixty-eight primipara women with expected due dates between May and December, 1992, volunteered to participate in the study. Of these, 34 were randomly assigned to each of the two groups. Approximately 81 percent of the women completed the study, leaving n = 29 in the control group and n = 26 in the intervention group. The intervention consisted of special incentives (prizes) for women and their partners to participate in several breast feeding education and promotion activities. Intervention group women and their partners experienced positive changes in breast feeding knowledge and attitudes. Furthermore, the intervention seemed to have influenced more women in the treatment group to breast feed despite their prenatal feeding intentions. In addition, the partners of intervention group women were perceived to be more supportive of breast feeding than control group partners. These findings suggest that incentives, such as donated prizes, can be used to attract lower socioeconomic group women and their partners to breast feeding promotion interventions. Participation in such interventions can produce positive changes in breast feeding knowledge, attitudes, and support, and can have a dramatic effect in promoting breast feeding.

Physical and functional S-layer reconstitution in Aeromonas salmonicida.

The various functions attributed to the S-layer of Aeromonas salmonicida have been previously identified by their conspicuous absence in S-layer-defective mutants. As a different approach to establish the multifunctional nature of this S-layer, we established methods for reconstitution of the S-layer of A. salmonicida. Then we investigated the functional competence of the reconstituted S-layer. S-layers were reconstituted in different systems: on inert membranes or immobilized lipopolysaccharide (LPS) from purified S-layer protein (A-protein) or on viable cells from either A-protein or preassembled S-layer sheets. In the absence of divalent cations and LPS, purified A-protein in solution spontaneously assembled into tetrameric oligomers and, upon concentration by ultrafiltration, into macroscopic, semicrystalline sheets formed by oligomers loosely organized in a tetragonal arrangement. In the presence of Ca2+, purified A-protein assembled into normal tetragonal arrays of interlocked subunits. A-protein bound with high affinity (Kd, 1.55 x 10(-7) M) and specificity to high-molecular-weight LPS from A. salmonicida but not to the LPSs of several other bacterial species. In vivo, A-protein could be reconstituted only on A. salmonicida cells which contained LPS, and Ca2+ affected both a regular tetragonal organization of the reattached A-protein and an enhanced reattachment of the A-protein to the cell surface. The reconstitution of preformed S-layer sheets (produced by an S-layer-secreting mutant) to an S-layer-negative mutant occurred consistently and efficiently when the two mutant strains were cocultured on calcium-replete solid media. Reattached A-protein (exposed on the surface of S-layer-negative mutants) was able to bind porphyrins and an S-layer-specific phage but largely lacked regular organization, as judged by its inability to bind immunoglobulins. Reattached S-layer sheets were regularly organized and imparted the properties of porphyrin binding, hydrophobicity, autoaggregation, adherence to and invasion of fish macrophages and epithelial cells, and resistance to macrophage cytotoxicity. However, cells with reconstituted S-layers were still sensitive to complement and insensitive to the antibiotics streptonigrin and chloramphenicol, indicating incomplete functional reconstitution.

Influences on breast-feeding by lower-income women: an incentive-based, partner-supported educational program.

OBJECTIVE: To determine the effects of a partner-supported, incentive-based educational program on rates and duration of breast-feeding among low-income women. DESIGN: Women who expressed a willingness to participate in the breast-feeding educational program were randomly assigned to one of two groups: an intervention group and a control group who received usual breast-feeding education. SETTING: Clinics of the Special Supplemental Food Program for Women, Infants, and Children in Flagstaff, Ariz. SUBJECTS: Sixty-eight primiparous pregnant women with expected due dates between May 1992 and December 1992 were willing to participate in the study. Of these, 34 were randomly assigned to the intervention group and 34 to the control group. Approximately 81% of the women completed the study: 29 in the control group and 26 in the intervention group. INTERVENTION: The intervention consisted of special incentives (prizes) for women and their partners to participate in a breast-feeding class for expectant couples and an educational series on childbirth. Women were also encouraged to use a breast-feeding support program in which peers serve as role models. MAIN OUTCOME MEASURES: The primary outcome measure was infant feeding method. Data were collected from mothers in both groups at the time of discharge from the hospital and at 2 weeks, 6 weeks, and 3 months postpartum. STATISTICAL ANALYSES PERFORMED: Binomial proportional analyses of the feeding data were performed. RESULTS: Women in the intervention group reported a higher percentage of breast-feeding at all measurement times. APPLICATIONS: These findings suggest that incentives, such as donated prizes, can be used to attract primiparous women from lower socioeconomic groups, along with their partners, to participate in educational interventions designed to promote breast-feeding. Participation by couples in breast-feeding promotion activities can dramatically increase the rate and duration of breast-feeding.

Physiological consequences of the S-layer of Aeromonas salmonicida in relation to growth, temperature, and outer membrane permeation.

S-layers are paracrystalline protein multimers that cover the entire cell surface of many bacterial species. The presence of an S-layer in Aeromonas salmonicida (also known as A-layer) predisposed this bacterium to apparently unrelated physiological consequences: inhibition of growth at 30 degrees C, enhanced cell filamentation at 37 degrees C, and enhanced uptake of the hydrophobic antibiotics streptonigrin and chloramphenicol. Growth inhibition or enhanced filamentation was not observed when the native A-layer was missing or its arrangement altered, as in Ca(2+)-limited or Ca(2+)- and Mg(2+)-limited cells, in A-layer-negative (A-) cells with an artificially reconstituted A-layer, or in mutants unable to correctly assemble this layer. A-layer-positive cells (A+) were far more sensitive to the intracellularly acting antibiotics streptonigrin and chloramphenicol than were A- cells, and streptonigrin-resistant mutants were predominantly A-. Hemin, a compound known to specifically bind to the A-layer, alleviated streptonigrin toxicity to A+, but not A-, cells. As well, Ca(2+)- and Mg(2+)-limited cells, or mutants harboring A-layer defects had a reduced sensitivity to streptonigrin, and A- cells with reconstituted A-layers remained resistant to streptonigrin and chloramphenicol. Thus, the presence of a native A-layer arrangement on the cell surface, and not the mere presence of A-layer protein subunits, predisposed A. salmonicida toward the aforementioned physiological consequences. The A-layer is suggested to specifically effect these consequences, in particular the permeation of streptonigrin or chloramphenicol, by a specific interaction of A-layer subunits with the outer membrane.

Novel structural patterns in divalent cation-depleted surface layers of Aeromonas salmonicida.

The fish pathogen Aeromonas salmonicida possesses a regular surface layer (or A-layer) which is an important virulence determinant. The A-protein, a single bilobed protein organized in a p4 lattice of M4C4 arrangement with two morphological domains, comprises this layer. The role of divalent cations in the A-layer structure was studied to better understand A-protein subunit interactions affecting structural flexibility and function. Divalent cation bridges were found to be involved in the integrity of the A-layer. Two novel A-layer patterns were formed as the result of growth under calcium limitation or by chelation of divalent cations with EDTA or EGTA, thereby constituting the first reported case of formation of distinct regular arrays upon divalent cation depletion. Furthermore, under these conditions A-protein was sometimes released as tetrameric units, rather than in monomeric form. The formation of the two novel patterns is best explained by a sequence of structural rearrangements, following disruption of only one of the two A-layer morphological units, that is, those held together by divalent cation bridges. The free tetrameric units represent four A-protein subunits clustered around the unaffected four-fold axis.

Structure of the tetragonal surface virulence array protein and gene of Aeromonas salmonicida.

The paracrystalline surface protein array of the pathogenic bacterium Aeromonas salmonicida is a primary virulence factor with novel binding capabilities. The species-specific structural gene (vapA) for this array protein (A-protein) was cloned into lambda gt11 but was unstable when expressed in Escherichia coli, undergoing an 816-base pair deletion due to a 21-base pair direct repeat within the gene. However, the gene was stable in cosmid pLA2917 as long as expression was poor. A-protein was located in the cytoplasmic, inner membrane and periplasmic fractions in E. coli. The DNA sequence revealed a 1,506-base pair open reading frame encoding a protein consisting of a 21-amino acid signal peptide, and a 481-residue 50,778 molecular weight protein containing considerable secondary structure. When assembled into a paracrystalline protein array on Aeromonas the cell surface A-protein was totally refractile to cleavage by trypsin, but became trypsin sensitive when disassembled. Trypsin cleavage of the isolated protein provided evidence that both the NH2- and COOH-terminal regions form distinct structural domains, consistent with three-dimensional ultrastructural evidence. The NH2-terminal 274-residue domain remained refractile to trypsin activity. This segment connects by a trypsin and CNBr-sensitive 78-residue linker region to a COOH-terminal 129-residue fragment which could apparently refold into a partially trypsin-resistant structure after cleavage at residue 323.

A novel ATPase complex selectively accumulated upon heat shock is a major cellular component of thermophilic archaebacteria.

We have discovered a large cylindrical protein complex which is an abundant component of the cytoplasm of extremely thermophilic archaebacteria. Structural analysis by image processing of electron micrographs suggests that the complex is composed of two stacked rings of eight subunits each; the rings enclose a central channel. The complex purified from the hyperthermophile Pyrodictium occultum is composed of equal quantities of two polypeptides of Mr 56,000 and 59,000. It exhibits an extremely thermostable ATPase activity with a temperature optimum of 100 degrees C. The basal level of the ATPase complex in the cell is high, and it becomes highly enriched as a result of heat shock (shift from 102 degrees C to 108 degrees C) or balanced growth at temperatures near the physiological upper limit. Immunoblotting results indicate that a related protein is present in most thermophilic archaebacteria and in Escherichia coli. This protein complex may play an important role in the adaptation of thermophilic archaebacteria to life at high temperature.